Functional self-association of von Willebrand factor during platelet adhesion under flow

Abstract

We have used recombinant wild-type human von Willebrand factor (VWF) and deletion mutants lacking the A1 and A3 domains, as well as specific function-blocking monoclonal antibodies, to demonstrate a functionally relevant self-association at the interface of soluble and surface-bound VWF. Platelets perfused at the wall shear rate of 1,500 s ؊1 over immobilized VWF lacking A1 domain function failed to become tethered to the surface when they were in a plasma-free suspension with erythrocytes, but adhered promptly if soluble VWF with functional A1 domain was added to the cells. The same results were observed when VWF was immobilized onto collagen through its A3 domain and soluble VWF with deleted A3 domain was added to the cells. Thus, VWF bound to glass or collagen sustains a process of homotypic self-association with soluble VWF multimers that, as a result, can mediate platelet adhesion. The latter finding demonstrates that direct immobilization on a substrate is not a strict requirement for VWF binding to platelet glycoprotein Ib␣. The dynamic and reversible interaction of surface-bound and soluble VWF appears to be specifically homotypic, because immobilized BSA, human fibrinogen, and fibronectin cannot substitute for VWF in the process. Our findings highlight a newly recognized role of circulating VWF in the initiation of platelet adhesion. The self-assembly of VWF multimers on an injured vascular surface may provide a relevant contribution to the arrest of flowing platelets opposing hemodynamic forces, thus facilitating subsequent thrombus growth. P latelet adhesion and aggregation at sites of vascular injury exposed to rapid blood flow, such as in arterioles of the normal circulation (1) or larger arteries with pathological lumen restrictions (2), require von Willebrand factor (VWF) (3-6). Under these hemodynamic conditions, immobilized VWF, either as an intrinsic subendothelial matrix component (7,8) or bound from plasma onto collagens (6, 9-11) and other extracellular substrates (12), mediates platelet deposition by interacting with the membrane glycoprotein (GP) Ib␣ receptor. Platelets become activated after the initial tethering, which allows their irreversible adhesion to the surface and the binding of plasma VWF and fibrinogen to the integrin ␣ IIb ␤ 3 . These ligands, immobilized on the membrane of adherent platelets, provide the substrate for the recruitment of additional platelets, which in turn become activated and bind the same plasma ligands repeating a cycle that supports continuing platelet-to-platelet cohesion, i.e., aggregation, into the thrombus mass (4).The domains of VWF involved in matrix and platelet interactions have been identified. Domain A3 plays a predominant role in VWF binding to collagen (13), and domain A1 interacts with platelet GP Ib␣ (14, 15), a component of the GP Ib-IX-V receptor complex (16). Finally, the C1 domain sequence ArgGly-Asp-Ser (RGDS), located near the C terminus of the VWF subunit (17,18), by binding to activated ␣ IIb ␤ 3 (19), stabilizes the initial platelet ...

ADAMTS-13 rapidly cleaves newly secreted ultralarge von Willebrand factor multimers on the endothelial surface under flowing conditions

“…25 This finding may represent the true length of a single, covalently bound multimer-which in molecular dimensions would rival some of the largest biologic molecules known-or alternatively could result from noncovalent self-association of VWF multimers, either in the Weibel-Palade bodies or after secretion, as has been demonstrated for surface-associated plasma forms of VWF. 26 It should be noted that we never observed the step-wise growth of the stringlike structures, as might be expected if additional multimers were being sequentially added from the newly released ULVWF. The difficulty in estimating the molecular masses of the largest multimeric forms of VWF by conventional means such as agarose gel electrophoresis renders estimation of ULVWF size problematic.…”

Force-induced on-rate switching and modulation by mutations in gain-of-function von Willebrand diseases

“…VWF circulates in the form of long, disulfidebonded concatemers, with tens to hundreds of monomers, which mostly adopt a compact, irregularly coiled conformation during normal hemodynamics (12). At sites of hemorrhage, flow changes from shear to elongational.…”

Kinetic study of von Willebrand factor self-aggregation induced by ristocetin

“…This process consists of a conformational change of VWF molecules, that expose within the A1 domain the binding site for GpIbα. This mechanism is triggered either by mechanical forces, such as high shear rate (>5000 sec -1 ) found in the arterial circulation, or by chemical potential generated by interaction with external surfaces and chemicals/snake venoms [5,[11][12][13][14][15]. Finally, natural mutations causing type 2B von Willebrand disease (VWD 2B) such as the Arg1306Trp or Arg1341Trp [6], are also able to stabilize a…”